The anti-dengue virus properties of statins may be associated with alterations in the cellular antiviral profile expression

Dengue virus (DENV) susceptibility to cholesterol depleting treatments has been previously reported. There are numerous questions regarding how DENV seizes cellular machinery and cholesterol to improve viral production and the effect of cholesterol sequestering agents on the cellular antiviral response. The aim of the present study was to evaluate the mechanisms involved in the negative regulation of DENV replication induced by agents that diminish intracellular cholesterol levels. Cholesterol synthesis was pharmacologically (fluvastatin, atorvastatin, lovastatin, pravastatin and simvastatin treatment) and genetically (HMGCR-RNAi) inhibited, in uninfected and DENV2-infected hepatoma Huh-7 cells. The cholesterol levels, DENV titer and cellular antiviral expression profile were evaluated. A reduction in the DENV titer, measured as plaque forming units, was observed in DENV-infected cells following 48 h treatment with 10 μM fluvastatin, 10 μM atorvastatin, 20 μM lovastatin and 20 μM simvastatin, which achieved 70, 70, 65 and 55% DENV2 inhibition, respectively, compared with the untreated cells. In addition, the cytopathic effect was reduced in the statin-treated DENV-infected cells. Statins simultaneously reduced cholesterol levels at 48 h, with the exception of DENV2 infected cells. Genetic inhibition of cholesterol synthesis was performed using RNA interference for 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR-siRNA), which indicated a slight reduction in DENV2 titer at 48 h post-infection, however, with no significant reduction in cholesterol levels. In addition, DENV2 infection was observed to augment the intracellular cholesterol levels in all experimental conditions. Comparison between the cellular antiviral response triggered by DENV2 infection, statin treatment and HMGCR-siRNA in infected, uninfected, treated and untreated Huh7 cells, showed different expression profiles for the antiviral genes evaluated. All downregulating cholesterol agents evaluated reduced the expression of genes associated with cellular immune and pro-inflammatory responses. These results indicate that statin-mediated downregulation of DENV2 infectious particles number is independent of cholesterol levels and it is partially mediated by the modulation of the cellular antiviral profile.